搜索结果：Critical reviews in biotechnology

Surface-enhanced Raman spectroscopy (SERS) is rapidly emerging as a transformative technology in dermatological diagnostics, offering ultra-sensitive, noninvasive detection of molecular markers associated with skin diseases. With nearly five billion individuals affected worldwide and conventional diagnostic methods often limited by invasiveness or subjective interpretation, there is an urgent need for rapid, accessible, and real-time diagnostic solutions. The present review systematically examines the integration of SERS into dermatological practice, with a focus on recent advancements in biosensing platforms, nanostructure engineering, and point-of-care devices. Innovative methodologies, including microneedle-based SERS biosensors and microfluidic-integrated detection systems, are discussed in the context of their ability to enhance diagnostic accuracy for early-stage skin cancers, microbial infections, and inflammatory dermatoses. Furthermore, the review highlighted the role of AI-driven spectral analysis in improving data interpretability and clinical decision-making. Critical evaluation of the challenges of substrate reproducibility, clinical standardization, and device scalability, while outlining emerging strategies that aim to bridge laboratory innovations with clinical applications, are explored. Looking ahead, the development of portable, low-cost SERS platforms for continuous skin health monitoring, combined with personalized diagnostic pathways, is poised to redefine dermatological care and expand the scope of precision medicine. Skin diseases impact 4.8 billion people globally, imposing substantial mortality and economic burdens, highlighting the urgent need for accurate, timely diagnostics. Current approaches are often invasive, time-intensive, or lack sensitivity. The present review critically examines the transformative potential of surface-enhanced Raman spectroscopy (SERS) as a minimally invasive, point-of-care solution for dermatological diseases and effective care management. The primary focus of this review is to explore the potential use of SERS in dermatological diagnostics, an area where traditional Raman spectroscopy has seen limited application. Integrating advanced spectroscopic techniques, lab-on-a-chip platforms and opto-microfluidic systems, it explores progressive SERS capabilities for early detection and disease management. By bridging medical engineering, medicine and biochemistry, this work addresses a critical knowledge gap, fostering interdisciplinary innovation to advance dermatological diagnostics and improve global health outcomes.

Fungi have emerged as powerful biological agents in the bioremediation of hydrocarbon-contaminated environments due to their robust enzymatic systems, adaptability, and ecological relevance. This review critically examines their potential, highlighting enzymatic mechanisms and technological innovations that underpin this sustainable approach. Case studies with: Aspergillus sp., Fusarium sp., Paecilomyces sp., Penicillium sp., and Trametes sp. demonstrate, through complex enzyme systems (laccases, lignin, manganese, and versatile peroxidases), the ability to degrade toxic compounds such as polycyclic aromatic hydrocarbons (PAHs) and BTEX, converting them into less harmful metabolites or even reusable by-products. Integrated strategies, including: biostimulation, bioaugmentation, microbial consortia, and the application of biosurfactants and bioemulsifiers, further enhance fungal efficiency in heterogeneous environments. Emerging innovations such as enzyme immobilization, myco-nanoremediation, and genetic engineering are discussed as promising solutions to overcome the inherent limitations of contaminant degradation under adverse conditions. Nevertheless, significant challenges remain, including the complexity of fungal metabolic pathways, gaps in proteomic regulation, and persistent obstacles in scaling and field reproducibility, which currently restrict large-scale commercial application. The alignment of fungal bioremediation with circular economy principles is emphasized, particularly the transformation of oil-derived pollutants into economically valuable bioproducts. Although fungal-based technologies remain underexplored commercially, especially in relation to regulatory frameworks and strategic partnerships, with this gap being especially evident in the Brazilian context, this review provides a robust foundation for advancing the application of fungi in sustainable environmental recovery. By integrating mechanistic insights with technological innovations and regulatory perspectives, it addresses a critical gap in the literature and outlines future directions for the field. This review uniquely integrates fungal bioremediation mechanisms with emerging technologies, highlighting enzyme immobilization and myco-nanoremediation. It addresses critical proteomic gaps that hinder scalability and field application, especially in hydrocarbon-polluted environments. It connects bioremediation to the principles of a circular economy by showcasing the various functions of fungi in breaking down hydrocarbons, absorbing heavy metals, and producing biosurfactants. Despite promising laboratory study results, it highlights the absence of commercial fungal technologies. It offers a pioneering outlook that bridges biochemical innovations and industrial deployment, especially within Brazil’s regulated framework, a perspective largely absent in recent global reviews.

Endophytic microorganisms are a vital part of the plant microbiome, contributing significantly to the plant's growth, development, and stress tolerance. Proteomics investigations have significantly enhanced our comprehension of the interactions between plants and endophytes, illuminating the complex molecular mechanisms that govern these mutually beneficial relationships. The review aims to integrate the latest developments in proteomic research concerning endophyte-plant interactions, emphasizing on elucidating the molecular mechanisms that underlie the benefit imparted to the host plant by the symbionts. The special focus of the review is to discuss the proteome level changes happening at the early recognition events, primary and secondary metabolism, signaling pathways, and defense mechanisms. By underscoring critical proteomic signatures, the review aspires to offer insights into how these interactions enhance plant health, increase stress resilience, and promote overall growth. The article discusses the potential applications of proteomics in agriculture and environmental sciences, emphasizing its role in crop resilience against biotic and abiotic stresses, optimizing biocontrol strategies, and improving nutrient use efficiency. The article also highlights that despite the advancements, critical gaps persist including the necessity for a deeper understanding of the temporal dynamics of proteomic responses, the specificity of protein-protein interactions, and the influence of environmental factors on the proteome induced by the endophytes. The review concludes by proposing future directions for proteomics research in plant-endophyte interactions for developing a more comprehensive understanding of the intricate molecular dialogues for developing a more sustainable and resilient agricultural systems. This review explores molecular interactions between plants and endophytic microorganisms particularly in context with proteomics research. It throws light on how endophytes enhance plant growth, stress tolerance and defense by modulating dynamic changes in protein expression during their association. Moreover, the review highlights practical applications of proteomics in optimizing biocontrol strategies, improving crop resilience, and increasing nutrient usage efficiency. Despite advancements, there remain significant knowledge gaps, emphasizing the need for further research to deepen our understanding of these molecular dialogues. Overall, the review integrates information on existing knowledge, potential future research directions, and emphasizes on the larger implications of such interactions in achieving sustainable agriculture and environmental outcomes.

Mass spectrometry (MS) has emerged as a powerful technique to study protein glycosylation. MS on intact denatured or native proteins can reveal all-inclusive glycoproteoform profiles while top-down, middle-down and/or bottom-up MS can uncover the characteristics of individual glycosylation sites. Alternatively, analyzing enzymatically released N-glycans can reveal intricate details on glycan isomers and generate high-throughput data on larger cohorts. All these methods are increasingly applied for the study of both individual glycoproteins and complex glycoprotein mixtures such as those originating from blood plasma or cell lysates. This has increased our knowledge about the complexity of protein glycosylation, but also revealed its huge diversity, which depends not only on the protein but also on the cell-dependent glycosylation machinery that may change with physiological conditions. Currently, multiple glycoproteins are recombinantly produced, for therapeutic applications as well as in the food sector, in host cells of diverse origin, most commonly: E. coli bacteria, yeast cells, insect cells, mammalian CHO or human HEK293 cells. Although glycoproteins of interest might show similar yields when produced in different host cells, an important question remains whether the host cell will or can provide similar or alike glycoproteoform profiles. In this review, we focus on the application of MS-based technologies to study glycosylation profiles of endogenous human glycoproteins and their recombinantly produced counterparts in different host cells. We will discuss in which ways recombinant glycoproteins can differ from their endogenous variants, and the functional consequences. Mass spectrometry (MS) is vital for uncovering protein glycosylation. This review investigates MS-based technologies used to explore the glycosylation profiles of human glycoproteins and their recombinant analogs. The emphasis is on various host cells for production, addressing discrepancies and their functional implications. Among other things, we describe that human plasma glycosylation is relatively simple and constrained, which is very different from that of recombinant proteins. While recombinant IgG1-based mAbs exhibit endogenous-like glycosylation traits, other proteins display major diversity in glycan size, linkage, and monosaccharide variations. This review enhances awareness of glycosylation complexity and the role of MS in glycoprotein analysis and production.

The global accumulation of keratin-rich waste, primarily from poultry and livestock industries, presents significant environmental and economic challenges. This review explores the potential of Bacillus-derived keratinases as a sustainable solution for keratin waste valorization and prospects of value-addition. Keratinases, the keratin hydrolyzing proteases produced predominantly by various Bacillus species, exhibit exceptional capability in degrading keratin, a highly stable and recalcitrant protein. This degradation process not only mitigates the environmental impact of keratin waste, but also converts it into valuable by-products with potential industrial applications. We systematically review various aspects, including: the production, properties and the mechanism of keratin degradation by Bacillus keratinases, highlighting their enzymatic properties, substrate specificity, and efficiency in valorizing keratin into peptides and amino acids. Biomolecular aspects and catalytic behavior relevant to the activity and stability of Bacillus keratinases are visited via in silico modeling. The economic and environmental benefits of utilizing keratinases for waste valorization are assessed, including reductions in waste disposal costs, greenhouse gas emissions, and the potential for creating new economic opportunities through the utilization of keratin-derived products. The recent advancements in keratin waste enzyme treatment and their utilization in developing circular bioeconomy are highlighted in the present article. Bacillus keratinases efficiently catalyze the breakdown of resilient keratin waste into value added productsSeveral Bacillus species are reported to be prolific producers of keratin hydrolyzing enzymes, hence it is important to understand their relatedness and differencesMolecular catalytic mechanism of the Bacillus keratinases delineates their functioningKeratinase based bioprocesses can be integrated into circular bioeconomy frameworks to optimize waste disposal and valorization.

Poly-γ-glutamic acid (γ-PGA) is a natural biopolymer with broad application potential. Molecular weight (MW) is a key physicochemical parameter governing its structural properties, functional performance, and application scope. Recently, sustainable biosynthesis of γ-PGA with controllable MW has gained attention because of its environmental sustainability and process flexibility. With advances in molecular editing and synthetic regulation, MW-control strategies have shifted from random mutagenesis-based strain improvement to precise gene-engineering regulation. This review surveys natural microbial resources producing γ-PGA with diverse MWs and engineering strategies enabling de novo γ-PGA biosynthesis in multiple microbial chassis. To address limited production efficiency, we summarize metabolic engineering approaches for improving γ-PGA yield and MW tunability, including precursor supply optimization, carbon-flux redistribution, transcriptional regulation, use of non-food renewable substrates, and mitigation of metabolic burden from multi-layered engineering. To promote customized production of low-MW γ-PGA, we highlight hydrolase-centered strategies, emphasizing hydrolase screening, optimization of expression elements, and coordinated regulation of γ-PGA stereochemical configuration. Finally, we review applications of γ-PGA with different MWs in food, biopharmaceutical, and agricultural sectors, critically examine links between molecular characteristics and application requirements, and discuss future functional diversification and industrial-scale development. The increasing scientific and industrial interest in γ-PGA stems from its versatile functional properties across agricultural, food, and industrial applications, in which MW is a key determinant of both biological activity and physicochemical behavior. This review systematically examines metabolic engineering strategies that enable the de novo design and customized biosynthesis of γ-PGA through coordinated modulation of synthase–hydrolase regulatory networks, within an integrated framework encompassing synthesis, modification, and application. The mechanistic insights presented here provide a rational roadmap and practical guidance for researchers advancing microbial biopolymer engineering.

As an economical and sustainable approach for the enhancement of oil recovery, microbial enhanced oil recovery has gained prominence in residual oil recovery from depleted oil reservoirs that are economically marginal or technically challenging due to low permeability, elevated water content and bearing heavy oil, particularly under current economic constraints in the petroleum industry. Over the last few decades, intensive efforts have been invested and extensive studies have been conducted regarding MEOR in both laboratory studies and field applications, with positive responses in oil reservoirs under different conditions. This study reviews recent advances in microbial enhanced oil recovery in oilfields in China, focusing on the latest knowledge regarding dominant microbial communities in oil reservoirs, microbes and their interactions in oil reservoirs and the key roles of microbial products in oil recovery. Strategies for microbial enhanced oil recovery and microbial enhanced energy recovery, including the novel concept of "Microbial plus" which refers to the microbial combined chemical method to enhance synergistic efficiency for enhanced oil recovery, are investigated. At last, recent field trials in Chinese oilfields were systematized based on the corresponding technologies utilized. This article highlights the strategic significance of microbial enhanced oil recovery (MEOR) in Chinese oilfields, emphasizing its feasibility in low-permeability, high-water-cut, and heavy-oil reservoirs. By identifying key microbial players and their metabolic roles, it clarifies the biological mechanisms of MEOR. Notably, the study introduces the innovative "Microbial plus" concept, integrating microbial and chemical methods to improve recovery efficiency. Field trial analyses further demonstrate the practical value of MEOR under current economic constraints. This work provides critical insights into sustainable oil recovery and offers a foundation for the broader application of MEOR technologies.

The increasing demand for sustainable sources of long chain polyunsaturated fatty acids (PUFAs), such as DHA and EPA, has driven research into alternative production methods. This review explores the potential of using FBW and by-products as substrates for microalgae cultivation, offering a cost-effective and environmentally friendly approach to PUFA production. Through a structured narrative and a deep bibliometric analysis, key trends and research hotspots were identified, highlighting the most promising microalgae and thraustochytrids species, including Aurantiochytrium sp., Phaeodactylum tricornutum, and Nannochloropsis oculata. These species demonstrated high lipid yields and significant PUFA content when grown on diverse FBW substrates, such as dairy by-products, molasses, and palm oil mill effluent. The review emphasizes the importance of pretreatment processes of recycled nutrients, such as enzymatic hydrolysis and fermentation, in enhancing nutrient bioavailability and optimizing microalgal growth. Economically, the use of FBW can reduce operating costs with potential increases in return on investment. However, challenges such as the initial setup costs of pretreatment processes and the need for contamination control must be addressed. To assist investors, a decision tree was developed, guiding through critical decision points, from resource assessment to process optimization and economic analysis. This tool supports informed decision-making, ensuring the balance of costs, benefits, and sustainability goals. This review assembles and maps the rapidly growing evidence on producing omega-3-rich lipids (DHA/EPA) from food-waste streams using microalgae and thraustochytrids. Combining a scoping review with bibliometric analysis, a global activity map was created, revealing key species-substrate pairings, and exposing blind spots in safety, standardization, and techno-economic issues. These insights were translated into an operational decision tree and metabolism-informed guidance for pretreatment and cultivation. Through reframing waste as a feedstock for high-value lipids, this work provides a clear evidence base and practical roadmap to accelerate sustainable biomanufacturing and inform policy, investment, and future research.

Integrating of immobilized enzymes into membrane technology represents a significant advancement in the field of membrane science, particularly in addressing issues of fouling and self-cleaning. This review explores the development, techniques, and applications of enzymatic membranes, focusing on their fouling-degrading and self-cleaning properties. Enzymatic membranes combine catalytic and separation functions, offering environmentally friendly solutions with high selectivity and mild operating conditions. The review traces the historical development from early attempts in the 1970s using proteases in polymeric membranes to recent advancements involving a variety of enzymes and support materials. Various immobilization techniques, such as covalent bonding and physical adsorption, are discussed in relation to their impact on membrane performance. Applications span multiple industries, including food, biofuels, and wastewater treatment. Despite significant progress, the field continues to evolve with emerging technologies and novel enzyme immobilization strategies, presenting ongoing opportunities for research and industrial application. This review intends to cover the last 50 years of publications on using immobilized enzymes to decrease membrane fouling and improve cleaning in membrane separation processes. By tracing the evolution of enzymatic membranes from their inception in the 1970s to recent advancements, it shows the rise of novel enzyme immobilization techniques and cutting-edge applications in biofuels production, wastewater treatment, and food processing. Enzyme immobilization on membranes represents an environmentally sustainable, highly selective, and efficient technology. This work not only consolidates existing knowledge but also identifies future research directions, underlining its critical contribution to advancing membrane science and technology.

Cultured meat production relies on the efficient: isolation, purification, expansion, and preservation of livestock-derived primary cells with robust proliferation and differentiation potential. Satellite cells and fibro-adipogenic progenitors from muscle and adipose tissues are key cellular sources for the in vitro reconstruction of meat-like structures. This review critically examines current methodologies used for primary cell processing in cultured meat scalable bioprocessing research. Tissue dissociation strategies are classified as enzymatic, non-enzymatic, and hybrid protocols and compared in terms of yield, scalability, and preservation of cell functionality. Cell purification techniques include: density gradient centrifugation, pre-plating, fluorescence-activated cell sorting (FACS), and magnetic-activated cell sorting (MACS), each offering different levels of specificity, throughput, and compatibility with food-grade applications. Long-term storage via cryopreservation is essential for establishing reliable cell banks. However, challenges remain in minimizing cryoinjury, selecting optimal cryoprotective agents, and maintaining post-thaw viability. Emerging alternatives, such as temperature-responsive cell sheet technology and biochemical protection strategies, are highlighted for their potential to improve cell recovery and functionality. Unlike existing reviews that often focus on isolated aspects, this article integrates dissociation, purification, and preservation into one framework. It highlights how optimized protocols can overcome current bottlenecks in cell processing. This review emphasizes the importance of standardized and food-safe platforms. Such advances will support commercialization and sustainable delivery of cultured meat products. This review addresses a critical challenge in cultured meat production: the lack of integrated, food-safe strategies for primary cell processing. Unlike previous reviews that examined cell isolation, sorting, or preservation in isolation, it unifies these steps into a single framework that balances biological performance with scalability and safety. By linking advanced cell handling technologies to reproducibility and commercial feasibility, the review provides actionable insights for both researchers and industry, supporting the development of sustainable and reliable cultured meat systems.

Pellets are ultrastructural configurations of filamentous fungal biomass that form during growth in submerged culture. This growth pattern offers advantages for controlling and stabilizing bioprocesses through biomass immobilization, reduced medium viscosity, and facilitated compound extraction. These benefits are particularly valuable for bioremediation, synergistic applications with biomaterials, and industrial metabolite production. However, fungal pellets also present challenges, such as limited oxygen diffusion to the pellet core, inconsistent pellet homogeneity, and decreased productivity. Factors such as electrostatic interactions, hydrophobicity, and culture conditions influence pellet formation. Currently, optimization efforts for pellet production focus on evaluating parameters, such as: pH range, agitation rate, pellet formation time, carbon source, additive agents, trace metals, and inoculum concentration, among others. Fungal pellets are increasingly recognized as promising platforms in emerging environmental biotechnology due to their versatility in pollutant removal and sustainable processing. Unlike previous reviews, this work provides an integrated and up-to-date perspective that combines the fundamentals of pellet formation with recent advances in their environmental and industrial applications, highlighting strategies for optimization and scalability. This review focuses on analyzing the most relevant aspects of fungal pellets, including their formation, optimization, and biotechnological applications. Given the growing importance of fungi in contemporary biotechnology, a state-of-the-art review of fungal pellets is warranted. This includes presenting an updated overview of processes for generating fungal biomass with enhanced handling, based on the use of fungal granules, an essential component for the implementation of efficient biotechnological processes using model fungal pellets with relevant industrial applications. This review critically examines the current and emerging applications of fungal pellets in biotechnology, highlighting their practical potential as well as their technical and operational limitations. While pellet formation is well known, their role as structured platforms for enzyme production, bioremediation, and metabolite synthesis demands updated evaluation. By integrating recent advances with unresolved challenges, this work provides a timely perspective that supports the rational design and implementation of fungal pellet-based processes across environmental and industrial contexts.

As an unconventional yeast, Yarrowia lipolytica is increasingly used in biomanufacturing high-value chemicals, including organic acids, sugar alcohols, fatty acid derivatives, and terpenes, owing to its unique physiological and biochemical properties. However, fine-tuning its biosynthetic pathways requires advanced transcriptional regulatory tools, with promoters being the most essential component. Numerous studies have sought to characterize promoter sequences by breaking them into distinct functional elements, while also constructing stronger and inducible synthetic promoters through rational design, modification, and optimization. This review summarizes the progress in discovering endogenous and engineering synthetic promoters in Y. lipolytica. We highlight emerging strategies such as hybrid promoter engineering, core promoter engineering, and transcription factor-based promoter engineering, which expand the promoter toolbox with enhanced properties. Finally, we address the current state of research and identify potential future directions in this field. This review highlights the latest advancements in promoter engineering for Yarrowia lipolytica, a promising microbial chassis for biomanufacturing high-value chemicals. By summarizing endogenous promoter discovery and synthetic promoter design strategies, we provide a comprehensive toolkit for fine-tuning metabolic pathways. Emerging approaches, such as hybrid and transcription factor-based promoter engineering, are discussed to address current limitations. This work not only advances the field of synthetic biology but also paves the way for more efficient and sustainable bioproduction processes, offering valuable insights for researchers and industry stakeholders.

The escalating problem of antibiotic resistance has sparked renewed interest in bacteriophages (phages) as potential substitutes for conventional antibiotics in treating infectious diseases, improving food safety, and advancing sustainable agriculture. The key phage research processes, such as host range, burst size, and environmental stability tests, strongly influence phage production processes. Hence, the standardization of the mentioned techniques must be prioritized. The introduction of high-throughput sequencing technologies with high accuracy and the emergence of novel bioinformatic tools to analyze the resulting raw molecular data provide comprehensive identification of phages and phage-verse (the universe of phage). While encapsulation of phages was studied comprehensively before, the production of encapsulated phages is still unclear. Moreover, recent advances in artificial intelligence (AI) contribute to phage research by increasing the accuracy of bioinformatic tools, improving resistance profiling, and facilitating phage host prediction. Incorporating AI promises a future of automated, precisely tailored phage applications. This review covers efficient techniques appropriate for industrial and agricultural applications as well as large-scale phage production methods, covering upstream and downstream processing. Also, encapsulated phage production and AI-based automated systems in various applications are proposed in this review. By covering both present issues and potential future uses of phages in the fight against antibiotic resistance, this review seeks to give academics and industry experts the fundamental information they need to advance phage-based solutions. Bacteriophages (phages) have been introduced as one of the most effective strategies to tackle antibiotic-resistant bacteria (superbugs). However, the lack of standard methods in phage investigation besides similarity makes published data unreliable in some cases. Also, this inconsistency affects phage production processes in industry. Notably, the existence of different formulas for calculating burst size can disrupt the proper choice of phages for antibacterial applications and make the simulation of the phage production process unreliable. This review comprehensively analyzes the most common and important techniques in phage research, highlighting gaps and proposed potential novel strategies from phage isolation to production.

The use of microalgal resources for bioplastic production has been hindered by resource flow limitations and large-scale production's economic viability. To address this limitation, this review intends to provide cost-effective methods for large-scale cultivation and surge in the accumulation of precursors for bioplastic production. The review shows how the engineering of metabolite-protein interaction favors optimization of growth conditions, nutrient availability, and harvesting techniques that increase the yield of bioplastic precursors from microalgae. The review shows that an in-depth understanding of metabolite-protein interaction for the biosynthesis of polyhydroxyalkanoates (PHA) can provide an important insight into the production of strains that will increase the bioplastic potential of microalgae. This research highlights the potential for optimizing metabolic pathways in microalgae to enhance the production of precursors linked to PHA, thus providing strategies that improve the efficiency and yield of these precursors from microalgae. The study also shows environmental stressors trigger post-translational modifications that enhance PHA production in microalgae. This post-translational change influences PHA synthesis by regulating metabolite-protein interaction that involves the activation of key enzymes in the biosynthetic pathway. Thus, indicating the benefit of manipulating these stressors to surge PHA accumulation in microalgae, thereby offering a sustainable solution for bioplastic production. This critical review proposes a pathway toward addressing global plastic pollution through sustainable production of PHA. The review deciphers enhancement of PHA productivity in microalgae, using efficient and environmentally friendly methods of plastic production, which is critical for enhancing cost-effectiveness and positive ecological footprints of biomanufacturing platforms.

L-Asparaginase (E.C.3.5.1.1; L-ASNase) hydrolyzes L-asparagine (an essential amino acid for the growth of leukemic cells) to aspartic acid and ammonia. It is obtained from various sources, including: bacteria, yeast, fungi, plants, and animals. It is used as a chemotherapeutic agent to treat acute lymphoblastic leukemia (ALL) and reduce acrylamide formation in baked and fried foods. Globally, various recombinant and pegylated L-ASNases formulations, including Escherichia coli and Erwinia-derived variants, are in Phase II or active clinical trials for ALL and related conditions across the USA, Brazil, and Canada. Some drugs are recruiting, and other variants of E. coli and E. chrysanthemi L-ASNases are already approved for marketing. Although L-ASNase is used as a therapeutic agent, immunogenic reactions and other adverse effects continue to limit its use. To enhance yield, L-ASNase isoforms are cloned and expressed in host cells, generating recombinants with distinct physicochemical properties and kinetic parameters. The enzymes' temperature and pH ranges vary from 25 to 100 °C and 6 to 10, respectively. Nowadays, enzymatic modifications, such as immobilization (chemical, physical, and PEGylation), mutagenesis, and PASylation, are used to overcome the limitations of commercialized L-ASNase-based drugs. Therefore, this review is a timely effort to compile and analyze the properties of recombinant L-ASNases and the contemporary techniques used to improve L-ASNase. A comprehensive study would help us better understand the kinetic parameters, biochemical properties, and modification trends of L-ASNase, enabling the development of robust, reliable therapeutics in the future. Currently available L-ASNase products often lack desirable pharmaceutical properties, including kinetic properties, increased half-life in blood serum, and decreased immunogenicity and toxicity. Several L-ASNases, including E. coli and Erwinia chrysanthemi, have been extensively characterized and evaluated in vitro for their anti-leukemic activity. However, only E. coli and Erwinia-derived formulations have advanced to preclinical animal models and subsequent clinical applications. Therefore, there is a critical need to identify new sources that are robust, more efficient, and have lower side effects. Hence, this review focuses on recombinant L-ASNases, their properties, drawbacks, and strategies for finding and improving L-ASNase variants.

Sugarcane, a leading source of sugar and bio-energy around the globe stands at the cross-road of genome complexity and agricultural innovation, offering the immense potential to fuel a sustainable future. Functional genomics with its precise identification and manipulation of genes could enable researchers unlock this potential and accelerate the breeding efforts. However, the polyploid genome of sugarcane with: high heterozygosity, high-repetitive DNA content, multiple copies of homo(eo)logous gene, epistatic interaction of alleles, etc., challenges the gene annotation, genome sequencing, genome editing, and phenotypic characterization. Similarly long breeding cycle, low transformation efficiency, time-consuming, and labor-intensive transformation methods further complicates the genome editing. Recent advances of functional genomics are transforming this scenario, such as current availability of reference genome "R570," has provided a significant insight of genome architect and function. Genome wide association studies (GWAS)/genome selection (GS) are enhancing trait-mapping and prediction of breeding values to accelerate the breeding cycles. The current era of smart breeding with integrative bio-informatics, advance genome editing tools, i.e., CRISPR/Cas-systems (Cas-proteins, Cas-RNPs, d-Cas-RNPs, and CRISPRa/i), and high-throughput phenomics offers a significant approach to: overcome transformation bottlenecks, explore complex trait architect and address polyploidy challenges. Therefore, this review summarizes the key challenges and focuses on elaborating recent advances and suggests optimized strategies for future improvement in functional genomics of sugarcane breeding. This review article gives a comprehensive overview of the question “Why sugarcane breeding is lagging behind in the current era of smart breeding?” and our opinions on “How can we address those reasons with recent advances in functional genomics?” Although various relevant articles (reference section) provide the significant insights into the functional identification and genome editing of sugarcane, but, novelty of our article is by focusing the key challenges, we provide efficient and transient functional genomics strategies in future breeding practices of sugarcane. Strategies suggested in this review will set path for many genome editing approaches of polyploidy crops in the future.

Enzymes are essential catalysts in numerous biological processes due to their efficiency, specificity, and selectivity. To enhance their stability and reusability, enzyme immobilization onto solid supports is crucial. Hierarchically porous metal-organic frameworks (HP-MOFs), with tunable meso- and macropores, have emerged as a promising solution for enzyme encapsulation, significantly improving catalytic performance. This review examines the development of various metal-based HP-MOFs, such as: iron, copper, zirconium, zinc, aluminum, and chromium, specifically designed for enzyme immobilization. The focus is on novel synthesis strategies, including functional group incorporation and hierarchical pore design, which optimize enzyme performance. Key advancements in immobilization techniques, such as: adsorption, covalent binding, in situ encapsulation, and post-synthetic infiltration, are also discussed. The review addresses the often-overlooked issue of HP-MOF toxicity, presenting both challenges and benefits. It also emphasizes the regulatory implications of HP-MOF applications, particularly in the food, pharmaceutical, and biomedical sectors, offering insights into how these materials can be safely integrated into these industries. Data from various studies demonstrate significant improvements in enzyme activity retention, stability, and efficiency in biocatalytic applications. Additionally, diverse applications in biocatalysis, biodiesel production, biosensing, and disease diagnosis are explored. A key feature of this review is the focus on advancing HP-MOF-enzyme composites toward higher technology readiness levels (TRLs), a topic not comprehensively covered in the literature. This review provides valuable insights for researchers aiming to optimize HP-MOFs for enzyme encapsulation and industrial biotechnological applications. This review presents a comprehensive analysis of hierarchically porous metal-organic frameworks (HP-MOFs) as next-generation platforms for enzyme immobilization. By integrating novel synthesis strategies with immobilization techniques, we highlight their potential in enhancing enzyme stability, activity, and reusability. Our focus on toxicity, regulatory considerations, and industrial applicability—especially regarding technology readiness levels (TRLs)—provides a unique perspective not previously addressed in the literature. This work aims to bridge fundamental research with real-world biotechnological applications, making it relevant for researchers, policymakers, and industry stakeholders alike.

Polymerase chain reaction (PCR) is a critical technology in nucleic acid detection and quantification. The PCR reaction requires thermal cycling the reaction mixture between two or more temperature stages for ∼30 cycles to achieve exponential amplification of the target DNA. Typically, the thermal cycling takes roughly an hour to finish and the large time consumption is a drawback for PCR. We review the various methods developed to reduce the thermal cycling time and build a rapid PCR. We group the methods to two approaches. The first approach is to increase the local heating/cooling power. The methods in this approach include contact heating, such as: heating resistors and Peltier pumps, and non-contact heating using air-blow, radiation on water and plasmonics. The other approach is to rapidly move the reaction mixture to a different temperature zone. Methods in this approach include: relocating the reaction vessel, continuous flow PCR using microfluidic chips, long tubes or oscillatory PCR scheme, and convective PCR. We analyze the advantages and challenges for each method used and the critical parameters to consider when evaluating the technologies. We review the technological advances and commercialization for each method. We also discuss the current challenges and future directions in building an effective and commercial rapid PCR, with the emphasis on sensitivity, portability and cost. A major drawback of polymerase chain reaction is its time consumption. Reducing the reaction time by performing rapid thermal cycling has been demonstrated using various methods. We analyze the advantages and disadvantages for each method used and discuss the critical parameters to consider in evaluating the technologies. We review the technological advancements in this field and introduce available commercial products. We also discuss the current challenges and future directions in building an effective rapid PCR. We aim to both inspire technological innovations and promote real-world commercial applications for rapid PCR.

Upon pathogen attack, cytosolic Ca2+ levels increase in plant cells. The first innate immune response is activated by detecting microbe/pathogen-associated molecular patterns (MAMPs/PAMPs) and is called PAMPs-triggered immunity (PTI). The second immune response is triggered by recognizing pathogens' effector proteins named effectors-triggered immunity (ETI). Calcium-dependent protein kinases (CDPKs or CPKs) are well-known calcium sensors that have a mediator role both in PTI and ETI. Calcium can bind to the elongation factor (EF)-hand domain at the C-terminus of CDPKs, which then phosphorylates substrates at the N-terminal catalytic kinase domain to transfer calcium signals directly. Improving the stress resilience of crops is a critical strategy in attaining global food security. In plants, when a stimulus is seen, there is an increase in Ca2+ concentration, which activates CDPKs which are in charge of sending out the immunological signals needed for disease tolerance. During the immune response, CDPKs are subject to numerous levels of regulation, including Ca2+ dependency to decipher various Ca2+ signals. Furthermore, salicylic acid (SA) regulation by CDPKs provides a comprehensive overview of CDPKs-mediated SA signaling during immune response in plants under pathogen attack. The critical part of CDPKs in SA biosynthesis, from the regulation of SA biosynthesis to how NPR1 perceives SA upon biotic stress, is comprehensively reviewed in this paper with the latest advancements in research. However, more research about CDPKs-mediated SA signaling under pathogen attack is mandatory to further dissect their co-role in crop protection against various diseases to achieve sustainable production goals in the future.